Conventional means for obtaining organic or protein molecules are often insufficient. Organic compounds are often extracted from biological materials (e.g., plants, microbes, and animals) or synthesized in the laboratory. Organic synthesis is usually complex since several steps are required to obtain the desired product. Furthermore, these steps often involve the use of toxic solvents, which require special handling and disposal. Extraction of organic compounds from biological materials may also require toxic solvents. In addition, extraction and purification methods usually provide a low yield of the desired compound, as biological materials typically contain only small quantities of these compounds.
Inducible promoter systems have been used to increase in vivo production of organic or protein molecules. Many of these systems, however, lack the ability to regulate the expression of the desired compound. For example, Saccharomyces cerevisiae inducible promoter systems have long been used for expression of heterologous proteins. Native inducible promoters such as GAL1 (Funk et al. (2002) Methods Enzymol. 350, 248-257), MET25 (Solow et al. (2005) Biotechnol. Prog. 21, 617-620), and CUP1 (Koller et al. (2000) Yeast 16, 651-656 and Mascorro-Gallardo et al. (1996) Gene 172, 169-170), although used successfully without modification, exhibit certain properties that are undesirable for the production of proteins and organic compounds. One common feature of these systems is their autocatalytic or switch-like behavior, where addition of small amounts of inducer leads to large increases in gene expression. In prokaryotes and bacteriophages, this is generally due to cooperative interactions between transcription factors and promoter elements. In more complex eukaryotic networks, other elements such as feedback loops, zero-order sensitivity, multi-step signaling mechanisms, and nucleocytoplasmic transport of regulatory proteins (Koshland (1998) Science 280, 852-853 and Verma et al. (2003) J. Biol. Chem. 278, 48764-48769) often contribute to nonlinear responses. In addition, native inducible promoter systems are often characterized by an all-or-none effect, in which genes are either maximally expressed or virtually not expressed in individual cells (Louis et al. (2002) Sci. STKE 2002, PE33). The inability to regulate gene expression in these inducible promoter systems can present problems such as toxicity, due to overproduction of the expressed compound. The inability to readily produce large quantities of many biological compounds has limited their practical use in areas such as drug production. Accordingly, there is a need for in vivo expression systems that can be tuned and regulated.